Examinando por Autor "Burdisso, Paula"
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Ítem Acceso Abierto Bacterially produced metabolites protect C. elegans neurons from degeneration(Public Library of Science, 2020-03-24) Urrutia, Arles; Garcia Angulo, Victor Antonio; Fuentes, Andrés; Caneo, Mauricio; Legüe, Marcela; Urquiza Zurich, Sebastian; Delgado, Scarlett E; Ugalde, Juan; Burdisso, Paula; Calixto, AndreaCaenorhabditis elegans and its cognate bacterial diet comprise a reliable, widespread model to study diet and microbiota effects on host physiology. Nonetheless, how diet influences the rate at which neurons die remains largely unknown. A number of models have been used in C. elegans as surrogates for neurodegeneration. One of these is a C. elegans strain expressing a neurotoxic allele of the mechanosensory abnormality protein 4 (MEC-4d) degenerin/epithelial Na+ (DEG/ENaC) channel, which causes the progressive degeneration of the touch receptor neurons (TRNs). Using this model, our study evaluated the effect of various dietary bacteria on neurodegeneration dynamics. Although degeneration of TRNs was steady and completed at adulthood in the strain routinely used for C. elegans maintenance (Escherichia coli OP50), it was significantly reduced in environmental and other laboratory bacterial strains. Strikingly, neuroprotection reached more than 40% in the E. coli HT115 strain. HT115 protection was long lasting well into old age of animals and was not restricted to the TRNs. Small amounts of HT115 on OP50 bacteria as well as UV-killed HT115 were still sufficient to produce neuroprotection. Early growth of worms in HT115 protected neurons from degeneration during later growth in OP50. HT115 diet promoted the nuclear translocation of DAF-16 (ortholog of the FOXO family of transcription factors), a phenomenon previously reported to underlie neuroprotection caused by down-regulation of the insulin receptor in this system. Moreover, a daf-16 loss-of-function mutation abolishes HT115-driven neuroprotection. Comparative genomics, transcriptomics, and metabolomics approaches pinpointed the neurotransmitter γ-aminobutyric acid (GABA) and lactate as metabolites differentially produced between E. coli HT115 and OP50. HT115 mutant lacking glutamate decarboxylase enzyme genes (gad), which catalyze the conversion of GABA from glutamate, lost the ability to produce GABA and also to stop neurodegeneration. Moreover, in situ GABA supplementation or heterologous expression of glutamate decarboxylase in E. coli OP50 conferred neuroprotective activity to this strain. Specific C. elegans GABA transporters and receptors were required for full HT115-mediated neuroprotection. Additionally, lactate supplementation also increased anterior ventral microtubule (AVM) neuron survival in OP50. Together, these results demonstrate that bacterially produced GABA and other metabolites exert an effect of neuroprotection in the host, highlighting the role of neuroactive compounds of the diet in nervous system homeostasis.Ítem Acceso Abierto Bases estructurales del reconocimiento ARN-proteína en el procesamiento de pequeños ARNs en plantas(Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas, 2014-03-06) Burdisso, Paula; Rasia, Rodolfo M.Los microARNs (miARNs) son moléculas de ARN pequeñas de 21 nucleótidos de longitud que se sintetizan en el núcleo por la ARN polimerasa II. En plantas, están involucrados en la regulación de procesos como el desarrollo, resistencia a estrés y respuestas a hormonas. La biogénesis de miARNs comienza con la transcripción de precursores mas largos, con extensa estructura secundaria de tallo y burbuja dentro de los cuales está contenida la secuencia que corresponde al mensaje de 21 nucleótidos. Estos precursores son procesados por un complejo proteico formado por la ARNasa III DICER-LIKE 1 (DCL1) y las proteínas accesorias HYPONASTIC LEAVES 1 (HYL1) y SERRATE (SE). Los precursores de plantas son sumamente heterogéneos. Sin embargo, la maquinaria de procesamiento, es capaz de liberar con precisión el miARN que posteriormente efectuará su acción regulando negativamente ARN mensajeros por complementariedad de bases de Watson y Crick. Durante los últimos años, se han dedicado muchos esfuerzos en descubrir nuevos miARNs, así como también se han logrado numerosos avances respecto a las formas de procesamiento de los precursores de plantas. Sin embargo, el mecanismo por el cual las proteínas de procesamiento llevan a cabo el reconocimiento es hasta el momento poco conocido. En este trabajo realizamos una caracterización biofísica de los dominios de unión a ARN doble hebra. En primer lugar se calculó la estructura en solución del segundo dsRBD de DCL1, a partir de la cual se observó que si bien tiene un plegamiento de dsRBD canónico, presenta diferencias con respecto a dominios homólogos. También se demostró que este dominio es capaz de unir tanto ARNdh como ADN, en contraste con lo que ocurre con la mayoría de los dsRBDs. La caracterización funcional de este dominio demostró que posiblemente actúe como una señal de localización atípica para direccionar a DCL1 al núcleo. Por otro lado, se analizaron los distintos determinantes de unión a sustrato del primer dsRBD de la proteína accesoria HYL1. Para esto, se generaron formas mutantes de la proteína, las cuales mantienen su estructura global, pero afectan las propiedades de unión al sustrato. Sorprendentemente, se demostró que una mutación y hasta una deleción completa en la región que se propone como principal determinante de unión al ARN no causa mayores efectos. Finalmente, se analizó la función de cada mutante in vivo, estableciendo una correlación directa entre la afinidad por los precursores y la actividad de la proteína.Ítem Acceso Abierto Induced folding in RNA recognition by Arabidopsis thaliana DCL1(Oxford University Press, 2015-06-22) Suárez, Irina Paula; Burdisso, Paula; Benoit, Matthieu P. M. H.; Boisbouvier, Jérôme; Rasia, Rodolfo M.Ítem Acceso Abierto Structural determinants of Arabidopsis thaliana Hyponastic Leaves 1 function in vivo(Public Library of Science, 2014-11-19) Burdisso, Paula; Milia, Fernando; Schapire, Arnaldo L.; Bologna, Nicolás G.; Palatnik, Javier F.; Rasia, Rodolfo M.MicroRNAs have turned out to be important regulators of gene expression. These molecules originate from longer transcripts that are processed by ribonuclease III (RNAse III) enzymes. Dicer proteins are essential RNAse III enzymes that are involved in the generation of microRNAs (miRNAs) and other small RNAs. The correct function of Dicer relies on the participation of accessory dsRNA binding proteins, the exact function of which is not well-understood so far. In plants, the double stranded RNA binding protein Hyponastic Leaves 1 (HYL1) helps Dicer Like protein (DCL1) to achieve an efficient and precise excision of the miRNAs from their primary precursors. Here we dissected the regions of HYL1 that are essential for its function in Arabidopsis thaliana plant model. We generated mutant forms of the protein that retain their structure but affect its RNA-binding properties. The mutant versions of HYL1 were studied both in vitro and in vivo, and we were able to identify essential aminoacids/residues for its activity. Remarkably, mutation and even ablation of one of the purportedly main RNA binding determinants does not give rise to any major disturbances in the function of the protein. We studied the function of the mutant forms in vivo, establishing a direct correlation between affinity for the pri-miRNA precursors and protein activity.Ítem Acceso Abierto The assembly of bacteria living in natural environments shapes neuronal integrity and behavioral outputs in Caenorhabditis elegans(American Society for Microbiology, 2023-04-25) Urquiza Zurich, Sebastian; Garcia Angulo, Victor Antonio; Burdisso, Paula; Palominos, M. Fernanda; Fernandez Hubeid, Lucia; Harcha, Paloma A.; Castillo, Juan P.; Calixto, Andrea; https://orcid.org/0000-0002-6168-7286Bacterivore nematodes are the most abundant animals in the biosphere, largely contributing to global biogeochemistry. Thus, the effects of environmental microbes on the nematodes’ life-history traits are likely to contribute to the general health of the biosphere. Caenorhabditis elegans is an excellent model to study the behavioral and physiological outputs of microbial diets. However, the effects of complex natural bacterial assemblies have only recently been reported, as most studies have been carried out with monoxenic cultures of laboratory-reared bacteria. Here, we quantified the physiological, phenotypic, and behavioral traits of C. elegans feeding on two bacteria that were coisolated with wild nematodes from a soil sample. These bacteria were identified as a putative novel species of Stenotrophomonas named Stenotrophomonas sp. strain Iso1 and a strain of Bacillus pumilus designated Iso2. The distinctive behaviors and developmental patterns observed in animals fed with individual isolates changed when bacteria were mixed. We studied in more depth the degeneration rate of the touch circuit of C. elegans and show that B. pumilus alone is protective, while the mix with Stenotrophomonas sp. is degenerative. The analysis of the metabolite contents of each isolate and their combination identified NAD1 as being potentially neuroprotective. In vivo supplementation shows that NAD1 restores neuroprotection to the mixes and also to individual nonprotective bacteria. Our results highlight the distinctive physiological effects of bacteria resembling native diets in a multicomponent scenario rather than using single isolates on nematodes.